Cellular biochemical reactions involving nucleic acids occur in aqueous solutions of salts, cosolutes, and biopolymers, such as proteins. These cosolutes, small organic solutes such as amino acids, nucleic acid precursors, simple sugars, and metabolites, can have dramatic influences on the structure and stability of nucleic acids. The long-term objective of this project is to elucidate the mechanism of cosolute-modulated folded nucleic acid stability, so as to better understand cosolute interactions with biopolymers and how these interactions influence biopolymer structural change and biochemical reactions. Additionally, these experiments will generate a cadre of undergraduate students at St. Olaf College that are trained to address biopolymer folding problems in a modern medical setting. The investigations detailed in this proposal will quantify accumulation or exclusion of glycine betaine, trimethylamine oxide (TMAO), and urea at nucleic acid surfaces to correlate cosolute interactions with chemical functional groups on the nucleic acid surfaces. The specific aims of this proposal will combine thermal unfolding and vapor pressure osmometry (VPO) studies with molecular dynamics (MD) computer simulations to: 1.) assess the strength of glycine betaine, TMAO, and urea as nucleic acid secondary structure stabilizers/destabilizers by quantifying the accumulation or exclusion of these cosolutes from chemical functional groups on double-helical DNA and RNA surfaces exposed during thermal denaturation;MD simulations will also be used to predict the roles solvent accessible chemical functional groups and base sequence-mediated hydration play in cosolute accumulation or exclusion at the double-helical DNA or RNA surface;2.) quantify the accumulation or exclusion of glycine betaine, TMAO, and urea from nucleoside 5'-monophosphates (NMPs), the individual building blocks of DNA and RNA secondary and tertiary structures, using VPO and MD simulations and couple these results with those from the first specific aim to elucidate the mechanism of cosolute stabilization or destabilization of DNA and RNA double-helices;3.) quantify the accumulation or exclusion of glycine betaine, TMAO, and urea from ribodinucleoside monophosphates (rDMPs) using VPO and MD simulations to assess the roles of base nearest-neighbor and stacking in cosolute interactions with DNA and RNA secondary and tertiary structures. These experiments will provide a foundation for an improved understanding of nucleic acid structural stability in cellular environments and a broader understanding of biopolymer folding and unfolding processes, leading to insights into biopolymer function and biopolymer folding diseases. PUBLIC HEALTH RELEVANCE: Secondary and tertiary structures of nucleic acids (DNA and RNA) are essential for proper biological function. This project seeks to understand how cellular solutes such as metabolites, amino acids, and sugars facilitate the gain or loss of nucleic acid structure. The experiments detailed in this proposal will improve our understanding of nucleic acid folding and unfolding processes and provide insights into nucleic acid function and biopolymer folding diseases.